Kinesin is a motor protein which uses the energy provided by ATP hydrolysis to move processively along microtubules. While two kinesin head domains are required for processive motion, the precise sequence of kinesin head-microtubule binding during ATP turnover is not known. Currently, the equilibrium models used to observe kinesin configuration at presumed turnover intermediates detect kinesin-microtubule binding only indirectly. The goal of this research is to directly determine the kinesin head binding status in the various model states, and the head binding status during ATP turnover. Whether the kinesin head is bound will be ascertained using fluorescence resonance energy transfer between a fluorescent dye attached to one head of a kinesin dimer, and a second fluorescent dye periodically and uniformly labeling a microtubule. Equilibrium experiments using ensembles of kinesin dimers will provide the population of bound and unbound heads under constant nucleotide conditions. Head binding during turnover will be directly determined using single kinesin dimers. Direct observation of binding at equilibrium and during turnover will help establish the relationship between the equilibrium models and actual turnover events.